WO2013058373A1 - Method for preventing deterioration of protein, and method for producing protein - Google Patents

Method for preventing deterioration of protein, and method for producing protein Download PDF

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Publication number
WO2013058373A1
WO2013058373A1 PCT/JP2012/077140 JP2012077140W WO2013058373A1 WO 2013058373 A1 WO2013058373 A1 WO 2013058373A1 JP 2012077140 W JP2012077140 W JP 2012077140W WO 2013058373 A1 WO2013058373 A1 WO 2013058373A1
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Prior art keywords
protein
packaging material
oxygen
storage container
atmosphere
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PCT/JP2012/077140
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French (fr)
Japanese (ja)
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貴幸 内田
智世至 石川
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ラジエ工業株式会社
協和発酵キリン株式会社
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Publication of WO2013058373A1 publication Critical patent/WO2013058373A1/en

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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B65—CONVEYING; PACKING; STORING; HANDLING THIN OR FILAMENTARY MATERIAL
    • B65B—MACHINES, APPARATUS OR DEVICES FOR, OR METHODS OF, PACKAGING ARTICLES OR MATERIALS; UNPACKING
    • B65B55/00—Preserving, protecting or purifying packages or package contents in association with packaging
    • B65B55/02—Sterilising, e.g. of complete packages
    • B65B55/04—Sterilising wrappers or receptacles prior to, or during, packaging
    • B65B55/08—Sterilising wrappers or receptacles prior to, or during, packaging by irradiation
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K41/00—Medicinal preparations obtained by treating materials with wave energy or particle radiation ; Therapies using these preparations
    • A61K41/10—Inactivation or decontamination of a medicinal preparation prior to administration to an animal or a person
    • A61K41/17—Inactivation or decontamination of a medicinal preparation prior to administration to an animal or a person by ultraviolet [UV] or infrared [IR] light, X-rays or gamma rays
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B65—CONVEYING; PACKING; STORING; HANDLING THIN OR FILAMENTARY MATERIAL
    • B65B—MACHINES, APPARATUS OR DEVICES FOR, OR METHODS OF, PACKAGING ARTICLES OR MATERIALS; UNPACKING
    • B65B31/00—Packaging articles or materials under special atmospheric or gaseous conditions; Adding propellants to aerosol containers
    • B65B31/02—Filling, closing, or filling and closing, containers or wrappers in chambers maintained under vacuum or superatmospheric pressure or containing a special atmosphere, e.g. of inert gas
    • B65B31/024—Filling, closing, or filling and closing, containers or wrappers in chambers maintained under vacuum or superatmospheric pressure or containing a special atmosphere, e.g. of inert gas specially adapted for wrappers or bags
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B65—CONVEYING; PACKING; STORING; HANDLING THIN OR FILAMENTARY MATERIAL
    • B65B—MACHINES, APPARATUS OR DEVICES FOR, OR METHODS OF, PACKAGING ARTICLES OR MATERIALS; UNPACKING
    • B65B55/00—Preserving, protecting or purifying packages or package contents in association with packaging
    • B65B55/02—Sterilising, e.g. of complete packages
    • B65B55/12—Sterilising contents prior to, or during, packaging
    • B65B55/16—Sterilising contents prior to, or during, packaging by irradiation
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B65—CONVEYING; PACKING; STORING; HANDLING THIN OR FILAMENTARY MATERIAL
    • B65B—MACHINES, APPARATUS OR DEVICES FOR, OR METHODS OF, PACKAGING ARTICLES OR MATERIALS; UNPACKING
    • B65B2220/00—Specific aspects of the packaging operation
    • B65B2220/16—Packaging contents into primary and secondary packaging
    • B65B2220/20—Packaging contents into primary and secondary packaging the primary packaging being bags, the secondary packaging being further bags, the primary bags being either finished or formed concurrently with the secondary bags

Definitions

  • the present invention relates to a method for preventing deterioration of a protein and a method for producing a protein using the method.
  • a storage container for a protein such as an antibody a sealable container made of glass or polymer has been used after being cleaned and sterilized using steam and / or a drug.
  • a sealable container made of glass or polymer has been used after being cleaned and sterilized using steam and / or a drug.
  • it can be very time consuming and expensive to establish that cleaning and sterilization is being performed reliably.
  • Patent Document 1 discloses a disposable processing bag having an alignment feature as an example of a single use bag.
  • Patent Document 2 discloses a method for manufacturing a prefilled syringe composed of polypropylene as a main material as a method for performing radiation sterilization in a deoxygenated atmosphere.
  • an object of the present invention is to provide a method for suppressing deterioration of a protein using a storage container sterilized by radiation for storing a protein solution.
  • the present inventors have found that the deterioration of the protein filled in the storage container can be suppressed by radiation sterilization of the storage container in a packaging material in a deoxygenated atmosphere, thereby completing the present invention.
  • the present invention is as follows. 1. A method for suppressing deterioration of a protein, characterized in that a storage container is subjected to radiation sterilization in a packaging material in a deoxygenated atmosphere. 2. 2. The method according to item 1 above, wherein an oxygen scavenger is enclosed in the packaging material to create a deoxidizing atmosphere in the packaging material. 3. 3. The method according to item 1 or 2 above, wherein an oxygen-absorbing material is used as the packaging material and the packaging material is deoxygenated. 4). A method for suppressing deterioration of a protein, characterized by radiation sterilizing a storage container in a packaging material enclosed with an oxygen scavenger. 5. 5. 5.
  • a storage container is radiation-sterilized in a packaging material in a deoxygenated atmosphere, whereby oxygen is activated by the radiation to become active oxygen, and the active oxygen is added to a component constituting the storage container or a protein solution. It is considered that it can prevent the components or additives from denatured and deteriorate the protein by acting on the product, and also prevent the active oxygen from acting on the protein and the protein from being deteriorated.
  • a method for suppressing deterioration of a protein a method for suppressing deterioration of a protein by the method, a method for producing a protein by suppressing deterioration of a protein by the method, a storage container capable of suppressing deterioration of the protein, and the storage container are used.
  • a method for producing a protein can be provided.
  • this method can also be applied to a method for sterilizing and suppressing deterioration of storage containers such as cells, DNA or RNA that are deteriorated by radiation.
  • FIG.1 (a) shows the perspective view of a single use bag.
  • FIG.1 (b) is a figure which shows the specific example of a single use bag.
  • FIG. 2 is a schematic diagram of a specific example in which protein degradation occurs due to radiation sterilization.
  • FIG. 3 is a plan view for explaining a method for sterilizing a single use bag.
  • FIG. 4 is a diagram showing the results of the example.
  • the present invention relates to a method for suppressing protein degradation, characterized in that a storage container is radiation sterilized in a packaging material in a deoxygenated atmosphere.
  • the “storage container” may be any one as long as it is composed of a polymer as a main material, and in particular may be one that can be stored by being filled with a liquid substance.
  • the inner surface (wetted surface) is preferably composed of a polymer, more preferably a single use bag, and even more preferably the liquid surface is composed of a polymer.
  • Polymer refers to various resin materials such as polyorphene or polyelastomer, preferably polyethylene, and more preferably ultra-low density polyethylene.
  • ultra-low density polyethylene refers to polyethylene having a specific gravity of 0.9 or less.
  • several different polymers can be used, for example, inflation method, T-die method, blow molding method, dry lamination method, hot-melt lamination method, co-extrusion inflation method, co-extrusion T-die method or hot press method. It may be laminated by various methods.
  • the resin material may contain various additives such as a plasticizer, an antioxidant, a flame retardant, a foaming agent, a color pigment, a filler, or a lubricant.
  • FIG. 1A shows a perspective view of a single use bag.
  • the single use bag 10 may be provided with a port 12, a tube 13, a stirring bar, or the like in the bag 11.
  • FIG. 1 (b) shows the style of a single use bag manufactured by Merck Millipore as a specific example of a single use bag.
  • Deoxygenated atmosphere refers to an atmosphere having a lower oxygen concentration than the atmospheric oxygen concentration (low oxygen atmosphere). Note that the oxygen concentration in the atmosphere is preferably as low as possible.
  • the method for obtaining a deoxygenated atmosphere include a method of reducing the oxygen concentration of the atmosphere using a deoxygenating agent.
  • a method for packaging a film or sheet having an oxygen-absorbing resin composition containing an oxidizing resin and a transition metal catalyst, or an oxygen-absorbing resin composition obtained by kneading the inorganic compound or organic compound in a resin A method for reducing the oxygen concentration of the atmosphere inside the packaging material by using it as a packaging material is also mentioned.
  • a method of filling the atmosphere with an inert gas such as argon or nitrogen Furthermore, for example, there is a method of reducing the oxygen from the atmosphere by reducing the pressure. These methods may be combined.
  • an inorganic system using iron powder, zinc powder, or an inorganic compound having an oxygen defect subjected to reduction treatment for example, titanium dioxide, zinc oxide, cerium oxide, iron oxide, etc.
  • An oxygen scavenger, an organic oxygen scavenger using glycerin, ascorbic acid or catechol, or an oxidizable resin for example, a thermoplastic resin having an ethylenically unsaturated bond, and a diamine component mainly composed of metaxylylenediamine
  • an oxygen-absorbing resin composition characterized by containing a transition metal catalyst and nylon MXD6 obtained by polycondensation from a dicarboxylic acid component containing adipic acid as a main component.
  • an oxygen-absorbing resin composition containing an oxidizing resin and a transition metal catalyst or an oxygen-absorbing resin composition obtained by kneading the inorganic compound or organic compound into a resin it can be processed into a film or sheet shape. These can be used as an oxygen scavenger after cutting into strips or the like. Further, the film or sheet is used as a component of a container such as a bag or case, or a multilayer container for packaging such as a bag or case (for example, for food packaging).
  • the “wrapping material” may be any material as long as it can package the storage container, and is preferably made of an oxygen-impermeable material.
  • a synthetic resin film or sheet laminated with an aluminum foil, an aluminum deposited film, or a synthetic resin film or sheet laminated with an oxygen barrier resin such as an ethylene-vinyl alcohol copolymer or vinylidene chloride is used. preferable.
  • an oxygen-absorbing resin composition containing an oxidizing resin and a transition metal catalyst, or a film or sheet having an oxygen-absorbing resin composition obtained by kneading the inorganic compound or organic compound into a resin An oxygen-absorbing material prepared using as a material may be used.
  • Random sterilization can use radiation normally used for radiation sterilization, and examples include gamma ( ⁇ ) rays, electron beams, and X-rays.
  • the radiation dose is also the dose used in normal radiation sterilization, and is not particularly limited, but is preferably 15 to 60 kilo gray, more preferably 25 to 45 kilo gray.
  • the irradiation time is not particularly limited, but is preferably 10 minutes to 7 hours.
  • Protein may be any protein. Specific examples include human serum proteins, peptide hormones, growth factors, cytokines, blood coagulation factors, fibrinolytic proteins, antibodies, and partial fragments of various proteins. Among these, a chimeric antibody, a monoclonal antibody such as a humanized antibody or a human antibody, an Fc fusion protein, an albumin-binding protein, or a partial fragment thereof is preferable. It should be noted that various substances such as cells, DNA or RNA that are denatured by active oxygen due to the denature mechanism can be prevented from being deteriorated by the method of the present invention.
  • Protein degradation means that physical and / or chemical changes such as polymerization, aggregation, degradation, deamidation, oxidation or conformational change are caused in the protein.
  • FIG. 2 is a schematic diagram showing a mechanism in which protein degradation occurs due to radiation sterilization of a storage container.
  • FIG. 2 shows a specific example where the protein is an antibody and the storage solution is a single use bag.
  • bag film composed of a polymer
  • oxygen contained in the atmosphere is excited by radiation, and reactive oxygen species Occurs.
  • the active oxygen causes denaturation of the protein contained in the filling of the storage container.
  • the active oxygen causes modification of the polymer constituting the single use bag and modification of the additive (for example, polysorbate 80) in the protein solution, and these modification components cause protein modification. It is thought that.
  • the method for suppressing protein degradation of the present invention can be applied to a method for producing a protein.
  • the “method for producing a protein” may be any method as long as it is a known method for producing a protein.
  • a storage container sterilized by radiation according to the method of the present invention can be used at the time of cell culture and when storing the drug substance after purification of the cell culture solution by various columns.
  • a deoxygenating atmosphere is created by encapsulating an oxygen scavenger in the packaging material, radiation sterilization is performed with gamma rays, and the antibody solution is filled into a single-use bag. This will be described as a representative example.
  • FIG. 3 is a plan view for explaining a method of sterilizing the single use bag 21.
  • a single use bag 21 having an inner surface (wetted surface) made of ultra-low density polyethylene is prepared.
  • the single use bag 21 may be attached to the bag 22 with an accessory such as a tube 23 or a port.
  • the single use bag 21 is packaged in the packaging material 24.
  • an oxygen scavenger sufficient to make the inside of the packaging material 24 a deoxygenated atmosphere is sealed.
  • the packaging material 24 is sealed, and the interior of the packaging material 24 is set to a deoxygenating atmosphere.
  • the packaging material 24 In order to seal the packaging material 24, heat fusion, high frequency fusion, fusion such as ultrasonic fusion, adhesion with an adhesive or a solvent, or the like can be used.
  • the packaging material 24 preferably has a gas barrier property that does not substantially transmit the atmosphere, and preferably does not shield radiation.
  • the oxygen scavenger 25 include iron-based oxygen scavengers and organic oxygen scavengers. 3)
  • the packaging material 24 is packaged in the packaging material 26. At the same time, an oxygen scavenger 25 sufficient to make the inside of the packaging material 26 a deoxygenating atmosphere is sealed.
  • the packaging material 26 is sealed using a method similar to the method for sealing the packaging material 24, and the interior of the packaging material 26 is set to a deoxygenated atmosphere.
  • the packaging material 26 preferably has the same characteristics as the packaging material 24. Radiation sterilization is performed by irradiating gamma rays from the outside of the packaging material 26. 5)
  • the packaging material 24 in which the sterilized packaging material 24 and the single use bag 21 are enclosed are opened in a clean atmosphere, and the packaging material 24 is taken out. Thereafter, the packaging material 24 is opened, and the single use bag 21 is taken out from the packaging material 24. 6) Fill the single use bag 21 with the antibody solution sterilized in advance. At that time, liquid is fed aseptically from the tube 23 or the port assembled to the bag 22.
  • the single-use bag 21 is subjected to radiation sterilization in a package in a deoxygenated atmosphere.
  • the packaging material 24 for the single-use bag 21 is used.
  • a method of enclosing the oxygen scavenger 25 inside and preferably also in the packaging material 26, a method of using a packaging material using an oxygen-absorbing material as the packaging material of the single-use bag 21, the packaging material 24 and preferably Examples include a method of filling the packaging material 26 with an inert gas, a method of reducing the pressure or vacuum in the packaging material 24, and preferably also in the packaging material 26.
  • the packaging material 24 and preferably the packaging material 26 are also preferably materials having gas barrier properties.
  • the present invention is not limited to this.
  • a single-use bag [Merck Millipore, FIG. 1 (b)] whose inner surface (wetted surface) was composed of ultra-low density polyethylene was prepared. Single-use bags are enclosed in double wrapping materials. An oxygen scavenger (manufactured by Mitsubishi Gas Chemical Company, “Ageless”) was enclosed in the packaging material, and the opening was sealed by heat sealing. As a comparison, a sample not containing an oxygen scavenger was also prepared.
  • radiation sterilization (gamma ray or electron beam, Radiage Industrial Co., Ltd.) was performed on the single use bag from the outside of the packaging material.
  • the radiation sterilization dose was 15, 30, 60 kg gray for the sample not containing the oxygen scavenger, and 60 kg gray for the sample containing the oxygen scavenger.
  • a specimen not subjected to radiation sterilization was also prepared as a control.
  • the specimen used for the SDS-PAGE test was subjected to analysis after cleaving the disulfide bond with a reducing reagent. Electrophoresis was performed by standard methods using an Invitrogen gel and electrophoresis apparatus. For staining, silver staining was performed using a silver staining kit manufactured by Cosmo Bio.
  • Figure 4 shows the results obtained. Since the degradation of the antibody appeared in a band around a molecular weight of 116 kilodaltons, the enlarged band is shown in the figure. Note that the darker the color of the band, the more advanced the deterioration.

Abstract

The purpose of the present invention is to provide a method for preventing the deterioration of a protein, in which a storage container sterilized by a radioactive ray is used for the storage of a solution of the protein. The present invention relates to: a method for preventing the deterioration of a protein, characterized by sterilizing a storage container by a radioactive ray in a packaging material in a deoxidized atmosphere; and a method for producing a protein, characterized by preventing the deterioration of the protein employing the above-mentioned method.

Description

たん白質の劣化を抑制する方法およびたん白質の製造方法Method for suppressing degradation of protein and method for producing protein
　本発明は、たん白質の劣化を防ぐ方法、および該方法を用いてたん白質を製造する方法に関する。 The present invention relates to a method for preventing deterioration of a protein and a method for producing a protein using the method.
　従来より、抗体等のたん白質の保存容器には、ガラスまたはポリマーで構成された密封可能な容器を洗浄後、蒸気および／または薬剤を使用し滅菌した後に使用していた。しかしながら、洗浄および滅菌が確実に行われていることを立証するには、多大な時間と費用がかかる。 Conventionally, as a storage container for a protein such as an antibody, a sealable container made of glass or polymer has been used after being cleaned and sterilized using steam and / or a drug. However, it can be very time consuming and expensive to establish that cleaning and sterilization is being performed reliably.
　したがって、抗体等の保存容器として１回使用限りのプラスチックバッグ（シングルユースバッグ）の利用が進められている。これらのバッグは滅菌が実施済みであり、洗浄を必要としないことから、使用者が要求される立証負担が軽減される。 Therefore, the use of a plastic bag (single-use bag) for one-time use as a storage container for antibodies and the like is being promoted. Since these bags have been sterilized and do not require cleaning, the burden of verification required by the user is reduced.
　特許文献１には、シングルユースバッグの一例として位置合わせフィーチャを有する使い捨て処理バッグが開示されている。また、特許文献２には、脱酸素雰囲気中で放射線滅菌を施す方法として、ポリプロピレンを主材料として構成されたプレフィルドシリンジの製造方法が開示されている。 Patent Document 1 discloses a disposable processing bag having an alignment feature as an example of a single use bag. Patent Document 2 discloses a method for manufacturing a prefilled syringe composed of polypropylene as a main material as a method for performing radiation sterilization in a deoxygenated atmosphere.
日本国特開２００８－２７３６２９号公報Japanese Unexamined Patent Publication No. 2008-273629 日本国特開２００４－２７５６１６号公報Japanese Unexamined Patent Publication No. 2004-275616
　本願発明者らは、公知の放射線滅菌方法により殺菌した保存容器をたん白質溶液の保存に用いた場合、その保存中にたん白質が劣化することを見出した。しかしながら、特許文献２においては、放射線により滅菌した保存容器をたん白質の保存に用いた場合のたん白質への影響について何ら検討はされていない。 The present inventors have found that when a storage container sterilized by a known radiation sterilization method is used for storing a protein solution, the protein deteriorates during the storage. However, in Patent Document 2, no consideration is given to the influence on the protein when a storage container sterilized by radiation is used for storing the protein.
　したがって、本発明は、放射線により滅菌した保存容器をたん白質溶液の保存に用いるたん白質の劣化を抑制する方法を提供することを目的とする。 Therefore, an object of the present invention is to provide a method for suppressing deterioration of a protein using a storage container sterilized by radiation for storing a protein solution.
　本発明者らは、保存容器を脱酸素雰囲気の包材中で放射線滅菌することにより、該保存容器に充填したたん白質の劣化を抑制できることを見出し、本発明を完成させた。 The present inventors have found that the deterioration of the protein filled in the storage container can be suppressed by radiation sterilization of the storage container in a packaging material in a deoxygenated atmosphere, thereby completing the present invention.
　すなわち、本発明は以下の通りである。
１．保存容器を脱酸素雰囲気の包材中で放射線滅菌することを特徴とするたん白質の劣化を抑制する方法。
２．前記包材中に脱酸素剤を同封することにより、該包材中を脱酸素雰囲気とする前項１に記載の方法。
３．前記包材として酸素吸入性素材を用いて該包材中を脱酸素雰囲気とする前項１または２に記載の方法。
４．脱酸素剤を同封した包材中で保存容器を放射線滅菌することを特徴とするたん白質の劣化を抑制する方法。
５．前記保存容器がポリマーで構成される前項１～４のいずれか１に記載の方法。
６．前記ポリマーがポリエチレンである前項５に記載の方法
７．前記ポリエチレンが超低密度ポリエチレンである前項６に記載の方法
８．前記保存容器がシングルユースバッグである前項１～７のいずれか１に記載の方法
９．前記放射線滅菌がγ線滅菌である前項１～８のいずれか１に記載の方法。
１０．前記たん白質が抗体である前項１～９のいずれか１に記載の方法。
１１．前記たん白質を含む溶液を前記放射線滅菌後の前記保存容器に充填して保存する前項１～１０のいずれか１に記載の方法。
１２．前項１～１１のいずれか１に記載の方法を用いてたん白質の劣化を抑制することを特徴とするたん白質を製造する方法。 That is, the present invention is as follows.
1. A method for suppressing deterioration of a protein, characterized in that a storage container is subjected to radiation sterilization in a packaging material in a deoxygenated atmosphere.
2. 2. The method according to item 1 above, wherein an oxygen scavenger is enclosed in the packaging material to create a deoxidizing atmosphere in the packaging material.
3. 3. The method according to item 1 or 2 above, wherein an oxygen-absorbing material is used as the packaging material and the packaging material is deoxygenated.
4). A method for suppressing deterioration of a protein, characterized by radiation sterilizing a storage container in a packaging material enclosed with an oxygen scavenger.
5. 5. The method according to any one of items 1 to 4, wherein the storage container is composed of a polymer.
6). 6. The method according to item 5 above, wherein the polymer is polyethylene. 7. The method according to item 6 above, wherein the polyethylene is ultra-low density polyethylene. 8. The method according to any one of items 1 to 7, wherein the storage container is a single use bag. 9. The method according to any one of items 1 to 8, wherein the radiation sterilization is γ-ray sterilization.
10. 10. The method according to any one of 1 to 9 above, wherein the protein is an antibody.
11. 11. The method according to any one of items 1 to 10, wherein the protein-containing solution is filled in the storage container after the radiation sterilization and stored.
12 12. A method for producing a protein, comprising suppressing deterioration of the protein using the method according to any one of 1 to 11 above.
　本発明は、保存容器を脱酸素雰囲気の包材中で放射線殺菌することにより、酸素が該放射線で活性化して活性酸素となり、該活性酸素が保存容器を構成する成分またはたん白質溶液中の添加物等に作用して該成分または添加物等が変性してたん白質を劣化させるのを防ぐとともに、活性酸素がたん白質に作用してたん白質を劣化させるのを防ぐことができると考えられる。 In the present invention, a storage container is radiation-sterilized in a packaging material in a deoxygenated atmosphere, whereby oxygen is activated by the radiation to become active oxygen, and the active oxygen is added to a component constituting the storage container or a protein solution. It is considered that it can prevent the components or additives from denatured and deteriorate the protein by acting on the product, and also prevent the active oxygen from acting on the protein and the protein from being deteriorated.
　本発明によれば、たん白質の劣化を抑制する方法、該方法によりたん白質の劣化を抑制してたん白質を製造する方法、たん白質の劣化が抑制可能な保存容器および該保存容器を用いてたん白質を製造する方法の提供が可能となる。また同様に、本方法は放射線によって劣化が生じる細胞、ＤＮＡまたはＲＮＡ等の保存容器の殺菌及び劣化の抑制方法にも応用可能である。 According to the present invention, a method for suppressing deterioration of a protein, a method for producing a protein by suppressing deterioration of a protein by the method, a storage container capable of suppressing deterioration of the protein, and the storage container are used. A method for producing a protein can be provided. Similarly, this method can also be applied to a method for sterilizing and suppressing deterioration of storage containers such as cells, DNA or RNA that are deteriorated by radiation.
図１（ａ）はシングルユースバッグの斜視図を示す。図１（ｂ）は、シングルユースバッグの具体例を示す図である。Fig.1 (a) shows the perspective view of a single use bag. FIG.1 (b) is a figure which shows the specific example of a single use bag. 図２は、放射線滅菌によりたん白質の劣化が生じる具体例の模式図である。FIG. 2 is a schematic diagram of a specific example in which protein degradation occurs due to radiation sterilization. 図３は、シングルユースバッグの滅菌方法を説明するための平面図である。FIG. 3 is a plan view for explaining a method for sterilizing a single use bag. 図４は、実施例の結果を示す図である。FIG. 4 is a diagram showing the results of the example.
　以下本発明を詳細に説明する。 Hereinafter, the present invention will be described in detail.
　本発明は、保存容器を脱酸素雰囲気の包材中で放射線滅菌することを特徴とするたん白質の劣化を抑制する方法に関する。 The present invention relates to a method for suppressing protein degradation, characterized in that a storage container is radiation sterilized in a packaging material in a deoxygenated atmosphere.
　「保存容器」とは、ポリマーを主材料で構成されるものであればいかなるものでもよく、特に液状の物質を充填して保存できるものであればよい。内表面（接液面）がポリマーで構成されたものであることが好ましく、シングルユースバッグがより好ましく、接液面がポリマーで構成されたものがさらに好ましい。 The “storage container” may be any one as long as it is composed of a polymer as a main material, and in particular may be one that can be stored by being filled with a liquid substance. The inner surface (wetted surface) is preferably composed of a polymer, more preferably a single use bag, and even more preferably the liquid surface is composed of a polymer.
　「ポリマー」とは、ポリオルフェンまたはポリエラストラマーなど種々の樹脂材料を指すが、好ましくはポリエチレンであり、より好ましくは超低密度ポリエチレンである。ここで、超低密度ポリエチレンとは、比重が０．９以下のポリエチレンをいう。また、ポリエチレンであれば数種の異なるポリマーを、例えば、インフレーション法、Ｔダイ法、ブロー成型法、ドライラミネート法、ホットメルトラミネート法、共押出インフレーション法、共押出Ｔダイ法またはホットプレス法等の種々の方法で積層させたものでもよい。前記樹脂材料中には、例えば可塑剤、酸化防止剤、難燃剤、発泡剤、着色顔料、充填剤または滑剤等の各種添加剤が含まれていてもよい。 “Polymer” refers to various resin materials such as polyorphene or polyelastomer, preferably polyethylene, and more preferably ultra-low density polyethylene. Here, ultra-low density polyethylene refers to polyethylene having a specific gravity of 0.9 or less. Further, in the case of polyethylene, several different polymers can be used, for example, inflation method, T-die method, blow molding method, dry lamination method, hot-melt lamination method, co-extrusion inflation method, co-extrusion T-die method or hot press method. It may be laminated by various methods. The resin material may contain various additives such as a plasticizer, an antioxidant, a flame retardant, a foaming agent, a color pigment, a filler, or a lubricant.
　「シングルユースバッグ」とは１回限りの使用を目的とした袋であり、溶液の調整若しくは保存、または細胞培養等の目的に使用される。溶液を無菌的に保存するためのシングルユースバッグは密封できるものが好ましい。図１（ａ）にシングルユースバッグの斜視図を示す。図１（ａ）に示すようにシングルユースバッグ１０には、バッグ１１にポート１２、チューブ１３または撹拌子などが付属していてもよい。図１（ｂ）にシングルユースバッグの具体例として、メルク・ミリポア社製シングルユースバッグの様式を示す。 “Single-use bags” are bags intended for one-time use, and are used for the purpose of preparing or storing solutions or cell culture. A single use bag for storing the solution aseptically is preferably sealable. FIG. 1A shows a perspective view of a single use bag. As shown in FIG. 1A, the single use bag 10 may be provided with a port 12, a tube 13, a stirring bar, or the like in the bag 11. FIG. 1 (b) shows the style of a single use bag manufactured by Merck Millipore as a specific example of a single use bag.
　「脱酸素雰囲気」とは、大気の酸素濃度よりも低い酸素濃度の雰囲気（低酸素雰囲気）のことを言う。なお、雰囲気中の酸素濃度は、できるだけ低い方が好ましい。脱酸素雰囲気にするための方法としては、例えば、脱酸素剤を用いて雰囲気の酸素濃度を低下させる方法が挙げられる。また、例えば、酸化性樹脂と遷移金属触媒とを含む酸素吸収性樹脂組成物または前記無機化合物もしくは有機化合物を樹脂に混練してなる酸素吸収性樹脂組成物を有するフィルムまたはシートを包装するための包材として用いることにより、その内部の雰囲気の酸素濃度を低下させる方法も挙げられる。 “Deoxygenated atmosphere” refers to an atmosphere having a lower oxygen concentration than the atmospheric oxygen concentration (low oxygen atmosphere). Note that the oxygen concentration in the atmosphere is preferably as low as possible. Examples of the method for obtaining a deoxygenated atmosphere include a method of reducing the oxygen concentration of the atmosphere using a deoxygenating agent. Further, for example, for packaging a film or sheet having an oxygen-absorbing resin composition containing an oxidizing resin and a transition metal catalyst, or an oxygen-absorbing resin composition obtained by kneading the inorganic compound or organic compound in a resin A method for reducing the oxygen concentration of the atmosphere inside the packaging material by using it as a packaging material is also mentioned.
　また、例えば、アルゴンまたはチッソ等の不活化ガスを雰囲気に満たす方法も挙げられる。さらに、例えば、減圧を行い雰囲気から酸素を減少させる方法も挙げられる。これらの方法を組み合わせてもよい。 Also, for example, a method of filling the atmosphere with an inert gas such as argon or nitrogen. Furthermore, for example, there is a method of reducing the oxygen from the atmosphere by reducing the pressure. These methods may be combined.
　「脱酸素剤」としては、例えば、鉄粉、亜鉛粉若しくは還元処理を施した酸素欠陥を有する無機化合物（例えば、二酸化チタン、酸化亜鉛、酸化セリウムまたは酸化鉄等）等を使用した無機系の脱酸素剤、グリセリン、アスコルビン酸若しくはカテコールを使用した有機系の脱酸素剤、または酸化性樹脂（例えば、エチレン系不飽和結合を有する熱可塑性樹脂、およびメタキシリレンジアミンを主成分とするジアミン成分とアジピン酸を主成分とするジカルボン酸成分から重縮合により得られるナイロンＭＸＤ６等）と遷移金属触媒とを含むことを特徴とする酸素吸収性樹脂組成物が挙げられる。これらの脱酸素剤は所定の酸素濃度、例えば食品保存のための所定の酸素濃度を達成できれば利用できる。 As the “deoxygenating agent”, for example, an inorganic system using iron powder, zinc powder, or an inorganic compound having an oxygen defect subjected to reduction treatment (for example, titanium dioxide, zinc oxide, cerium oxide, iron oxide, etc.) An oxygen scavenger, an organic oxygen scavenger using glycerin, ascorbic acid or catechol, or an oxidizable resin (for example, a thermoplastic resin having an ethylenically unsaturated bond, and a diamine component mainly composed of metaxylylenediamine) And an oxygen-absorbing resin composition characterized by containing a transition metal catalyst and nylon MXD6 obtained by polycondensation from a dicarboxylic acid component containing adipic acid as a main component. These oxygen scavengers can be used if they can achieve a predetermined oxygen concentration, for example, a predetermined oxygen concentration for food preservation.
　酸化性樹脂と遷移金属触媒とを含む酸素吸収性樹脂組成物または前記無機化合物もしくは有機化合物を樹脂に混練してなる酸素吸収性樹脂組成物の場合にはフィルムまたはシートの形状に加工できるので、これらを短冊状等に切断後、脱酸素剤として利用することができる。さらに前記のフィルムまたはシートは、袋若しくはケースなどの容器、または袋若しくはケースなどの包装用（例えば、食品包装用）多層容器の構成成分として使用される。 In the case of an oxygen-absorbing resin composition containing an oxidizing resin and a transition metal catalyst or an oxygen-absorbing resin composition obtained by kneading the inorganic compound or organic compound into a resin, it can be processed into a film or sheet shape. These can be used as an oxygen scavenger after cutting into strips or the like. Further, the film or sheet is used as a component of a container such as a bag or case, or a multilayer container for packaging such as a bag or case (for example, for food packaging).
　「包材」とは保存容器を包装できるものであればどのようなものでもよく、好ましくは酸素不透過性の材料から作成されたものがよい。材料としては、特に、合成樹脂フィルムまたはシートにアルミニウム箔を積層したもの、アルミニウム蒸着したもの、あるいはエチレン―ビニルアルコール共重合体若しくは塩化ビニリデンなどの酸素バリアー性樹脂を積層した合成樹脂フィルムまたはシートが好ましい。 The “wrapping material” may be any material as long as it can package the storage container, and is preferably made of an oxygen-impermeable material. As the material, in particular, a synthetic resin film or sheet laminated with an aluminum foil, an aluminum deposited film, or a synthetic resin film or sheet laminated with an oxygen barrier resin such as an ethylene-vinyl alcohol copolymer or vinylidene chloride is used. preferable.
　また、包材の材料としては、酸化性樹脂と遷移金属触媒とを含む酸素吸収性樹脂組成物または前記無機化合物もしくは有機化合物を樹脂に混練してなる酸素吸収性樹脂組成物を有するフィルムまたはシートを材料として作成された酸素吸収性素材でもよい。 Further, as a material of the packaging material, an oxygen-absorbing resin composition containing an oxidizing resin and a transition metal catalyst, or a film or sheet having an oxygen-absorbing resin composition obtained by kneading the inorganic compound or organic compound into a resin An oxygen-absorbing material prepared using as a material may be used.
　「放射線滅菌」とは放射線滅菌に通常用いられる放射線を用いることができ、例えば、ガンマ（γ）線、電子線およびＸ線が挙げられる。放射線の線量も、通常の放射線滅菌で用いられる線量が用いられ、特に限定されないが、１５～６０キログレイが好ましく、２５～４５キログレイがより好ましい。また、放射線照射時間は、特に限定されないが、１０分～７時間が好ましい。 “Radiation sterilization” can use radiation normally used for radiation sterilization, and examples include gamma (γ) rays, electron beams, and X-rays. The radiation dose is also the dose used in normal radiation sterilization, and is not particularly limited, but is preferably 15 to 60 kilo gray, more preferably 25 to 45 kilo gray. The irradiation time is not particularly limited, but is preferably 10 minutes to 7 hours.
　「たん白質」とはいかなるたん白質でもよい。具体的には、例えば、ヒト血清たん白質、ペプチドホルモン、増殖因子、サイトカイン、血液凝固因子、線溶系たん白質、抗体および各種たん白質の部分断片などが挙げられる。これらの中でも、キメラ抗体、ヒト化抗体若しくはヒト抗体などのモノクローナル抗体、Ｆｃ融合たん白質、アルブミン結合たん白質または該部分断片などが好ましい。なお、変性メカニズムから、活性酸素により変性が引き起こされる、細胞、ＤＮＡまたはＲＮＡなどの種々物質も本発明の方法により劣化を防ぐことができる。 “Protein” may be any protein. Specific examples include human serum proteins, peptide hormones, growth factors, cytokines, blood coagulation factors, fibrinolytic proteins, antibodies, and partial fragments of various proteins. Among these, a chimeric antibody, a monoclonal antibody such as a humanized antibody or a human antibody, an Fc fusion protein, an albumin-binding protein, or a partial fragment thereof is preferable. It should be noted that various substances such as cells, DNA or RNA that are denatured by active oxygen due to the denature mechanism can be prevented from being deteriorated by the method of the present invention.
　「たん白質の劣化」とは、重合、凝集、分解、脱アミド、酸化またはコンフォメーション変化などの物理的および／または化学的変化がたん白質に引き起こされることをいう。 “Protein degradation” means that physical and / or chemical changes such as polymerization, aggregation, degradation, deamidation, oxidation or conformational change are caused in the protein.
　図２は、保存容器の放射線滅菌によりたん白質の劣化が生じるメカニズムを示す模式図である。図２では、具体例として、たん白質が抗体であり、保存溶液がシングルユースバッグである場合を示す。図２に示すように、ポリマーで構成されたシングルユースバッグのフィルム（バッグフィルム）に対して、大気中で放射線滅菌を施した場合、大気中に含まれる酸素が放射線により励起され、活性酸素種が生じる。該活性酸素が、保存容器の充填物に含まれるたんぱく質の変性を引き起こす。また、該活性酸素によりシングルユースバッグを構成するポリマーの変性が生じると共に、たん白質溶液中の添加剤（例えば、ポリソルベート８０等）の変性が生じ、これらの変性成分によってもたん白質の変性が引き起こされると考えられる。 FIG. 2 is a schematic diagram showing a mechanism in which protein degradation occurs due to radiation sterilization of a storage container. FIG. 2 shows a specific example where the protein is an antibody and the storage solution is a single use bag. As shown in FIG. 2, when radiation sterilization is performed in the atmosphere on a single-use bag film (bag film) composed of a polymer, oxygen contained in the atmosphere is excited by radiation, and reactive oxygen species Occurs. The active oxygen causes denaturation of the protein contained in the filling of the storage container. In addition, the active oxygen causes modification of the polymer constituting the single use bag and modification of the additive (for example, polysorbate 80) in the protein solution, and these modification components cause protein modification. It is thought that.
　本発明のたん白質の劣化を抑制する方法は、たん白質を製造する方法に適用することができる。「たん白質を製造する方法」とはたん白質を製造する公知の方法であればいずれの方法でもよい。たん白質を製造する公知の方法において、細胞培養時及び各種カラムによる細胞培養液の精製後の原薬保存時等に本発明の方法で放射線殺菌した保存容器を用いることが出来る。 The method for suppressing protein degradation of the present invention can be applied to a method for producing a protein. The “method for producing a protein” may be any method as long as it is a known method for producing a protein. In a known method for producing a protein, a storage container sterilized by radiation according to the method of the present invention can be used at the time of cell culture and when storing the drug substance after purification of the cell culture solution by various columns.
　超低密度ポリエチレンで構成されるシングルユースバッグに対して、包材内に脱酸素剤を封入することにより脱酸素雰囲気とし、ガンマ線により放射線滅菌を施し、抗体溶液をシングルユースバッグに充填した場合を代表例として説明する。 For single-use bags made of ultra-low-density polyethylene, a deoxygenating atmosphere is created by encapsulating an oxygen scavenger in the packaging material, radiation sterilization is performed with gamma rays, and the antibody solution is filled into a single-use bag. This will be described as a representative example.
　図３はシングルユースバッグ２１の滅菌方法を説明するための平面図である。
１）まず、内表面（接液面）が超低密度ポリエチレンで構成されたシングルユースバッグ２１を用意する。シングルユースバッグ２１は図３に示すように、バッグ２２にチューブ２３またはポートなどの付属品が組みつけられていてもよい。
２）次に、シングルユースバッグ２１を包材２４内に包装する。同時に包材２４内を脱酸素雰囲気とするのに十分な脱酸素剤を封入する。シングルユースバッグ２１および脱酸素剤２５を封入したのち、包材２４を密閉し、包材２４の内部を脱酸素雰囲気とする。包材２４を密封するために、熱融着、高周波融着、超音波融着のような融着、または接着剤若しくは溶媒による接着等を用いることができる。包材２４は大気を実質的に透過させないガスバリア性を有するものが好ましく、また、放射線を遮蔽しないものが好ましい。脱酸素剤２５としては、例えば、鉄系の脱酸素剤および有機系の脱酸素剤等が挙げられる。
３）さらに、包材２６内に包材２４を包装する。同時に包材２６内を脱酸素雰囲気とするのに十分な脱酸素剤２５を封入する。包材２４および脱酸素剤２５を同封したのち、上記包材２４を密封するのと同様の方法を用いて包材２６を密閉し、包材２６の内部を脱酸素雰囲気とする。包材２６は包材２４と同様の特徴を有するものが好ましい。包材２６の外部からガンマ線を照射して放射線滅菌を施す。
５）滅菌が完了した包材２４およびシングルユースバッグ２１が同封された包材２６を清浄な雰囲気下で開封し、包材２４を取り出す。その後、包材２４を開封し、包材２４からシングルユースバッグ２１を取り出す。
６）あらかじめ滅菌処理された抗体溶液をシングルユースバッグ２１に充填する。その際、バッグ２２に組みつけられたチューブ２３またはポートから無菌的に送液を行う。 FIG. 3 is a plan view for explaining a method of sterilizing the single use bag 21.
1) First, a single use bag 21 having an inner surface (wetted surface) made of ultra-low density polyethylene is prepared. As shown in FIG. 3, the single use bag 21 may be attached to the bag 22 with an accessory such as a tube 23 or a port.
2) Next, the single use bag 21 is packaged in the packaging material 24. At the same time, an oxygen scavenger sufficient to make the inside of the packaging material 24 a deoxygenated atmosphere is sealed. After enclosing the single-use bag 21 and the oxygen scavenger 25, the packaging material 24 is sealed, and the interior of the packaging material 24 is set to a deoxygenating atmosphere. In order to seal the packaging material 24, heat fusion, high frequency fusion, fusion such as ultrasonic fusion, adhesion with an adhesive or a solvent, or the like can be used. The packaging material 24 preferably has a gas barrier property that does not substantially transmit the atmosphere, and preferably does not shield radiation. Examples of the oxygen scavenger 25 include iron-based oxygen scavengers and organic oxygen scavengers.
3) Further, the packaging material 24 is packaged in the packaging material 26. At the same time, an oxygen scavenger 25 sufficient to make the inside of the packaging material 26 a deoxygenating atmosphere is sealed. After enclosing the packaging material 24 and the oxygen scavenger 25, the packaging material 26 is sealed using a method similar to the method for sealing the packaging material 24, and the interior of the packaging material 26 is set to a deoxygenated atmosphere. The packaging material 26 preferably has the same characteristics as the packaging material 24. Radiation sterilization is performed by irradiating gamma rays from the outside of the packaging material 26.
5) The packaging material 24 in which the sterilized packaging material 24 and the single use bag 21 are enclosed are opened in a clean atmosphere, and the packaging material 24 is taken out. Thereafter, the packaging material 24 is opened, and the single use bag 21 is taken out from the packaging material 24.
6) Fill the single use bag 21 with the antibody solution sterilized in advance. At that time, liquid is fed aseptically from the tube 23 or the port assembled to the bag 22.
　本発明の方法においては、シングルユースバッグ２１に対して脱酸素雰囲気の包装中で放射線滅菌を施すものであり、脱酸素雰囲気を作り出す方法として、例えば、シングルユースバッグ２１に対して、包材２４内および、好ましくは包材２６内においても脱酸素剤２５を同封する方法、シングルユースバッグ２１の包材として酸素吸収性素材を用いた包材を使用する方法、包材２４内および、好ましくは包材２６内においても不活性ガスにより満たす方法、包材２４内および、好ましくは包材２６内においても減圧または真空状態とする方法が挙げられる。これらの場合、包材２４および、好ましくは包材２６においてもガスバリア性を有する素材が好ましい。 In the method of the present invention, the single-use bag 21 is subjected to radiation sterilization in a package in a deoxygenated atmosphere. As a method for creating a deoxygenated atmosphere, for example, the packaging material 24 for the single-use bag 21 is used. A method of enclosing the oxygen scavenger 25 inside and preferably also in the packaging material 26, a method of using a packaging material using an oxygen-absorbing material as the packaging material of the single-use bag 21, the packaging material 24 and preferably Examples include a method of filling the packaging material 26 with an inert gas, a method of reducing the pressure or vacuum in the packaging material 24, and preferably also in the packaging material 26. In these cases, the packaging material 24 and preferably the packaging material 26 are also preferably materials having gas barrier properties.
　以上、本発明のたん白質の劣化を抑制する方法およびたん白質の製造方法に関して説明したが、本発明はこれに限定されるものではない。 As mentioned above, although the method for suppressing the deterioration of the protein and the method for producing the protein of the present invention have been described, the present invention is not limited to this.
シングルユースバッグの滅菌
　内表面（接液面）が超低密度ポリエチレンで構成されたシングルユースバッグ［メルク・ミリポア社製、図１（ｂ）］を準備した。シングルユースバッグは２重の包材中に封入されている。その包材中に脱酸素剤（三菱ガス化学社製、「エージレス」）を同封し、開口部を熱融着により封止した。比較として、脱酸素剤を同封していない検体も準備した。 Single-use bag sterilization A single-use bag [Merck Millipore, FIG. 1 (b)] whose inner surface (wetted surface) was composed of ultra-low density polyethylene was prepared. Single-use bags are enclosed in double wrapping materials. An oxygen scavenger (manufactured by Mitsubishi Gas Chemical Company, “Ageless”) was enclosed in the packaging material, and the opening was sealed by heat sealing. As a comparison, a sample not containing an oxygen scavenger was also prepared.
　次に、包材の外部からシングルユースバッグに対して、放射線滅菌（ガンマ線または電子線、ラジエ工業社）を実施した。なお、放射線滅菌の線量は脱酸素剤を同封していない検体に対して１５，３０，６０キログレイとし、脱酸素剤を同封した検体は６０キログレイとした。また、対照として放射線滅菌を施していない検体も準備した。 Next, radiation sterilization (gamma ray or electron beam, Radiage Industrial Co., Ltd.) was performed on the single use bag from the outside of the packaging material. The radiation sterilization dose was 15, 30, 60 kg gray for the sample not containing the oxygen scavenger, and 60 kg gray for the sample containing the oxygen scavenger. In addition, a specimen not subjected to radiation sterilization was also prepared as a control.
抗体溶液への影響評価
　「シングルユースバッグの滅菌」で準備したシングルユースバッグにろ過滅菌により滅菌された抗体溶液を充填し、２５℃に温度管理されたインキュベータに１カ月保存した後、抗体の劣化をＳＤＳ－ＰＡＧＥ試験法により評価を行った。 Assessment of effects on antibody solutions Single-use bags prepared in "Sterilization of single-use bags" are filled with antibody solutions sterilized by filtration sterilization and stored in an incubator controlled at 25 ° C for one month, followed by antibody degradation. Was evaluated by the SDS-PAGE test method.
　ＳＤＳ－ＰＡＧＥ試験に用いる検体は、還元試薬を用いてジスルフィド結合を切断したのち分析に供した。泳動はインビトロジェン社のゲルおよび泳動装置を用いて標準的な方法で実施した。染色はコスモバイオ社製の銀染キットを用いて銀染色を実施した。 The specimen used for the SDS-PAGE test was subjected to analysis after cleaving the disulfide bond with a reducing reagent. Electrophoresis was performed by standard methods using an Invitrogen gel and electrophoresis apparatus. For staining, silver staining was performed using a silver staining kit manufactured by Cosmo Bio.
　図４に得られた結果を示す。抗体の劣化は分子量１１６キロダルトン付近のバンドに表れていたため、そのバンドを拡大したものを図示した。なお、バンドの色が濃いほど劣化が進んでいることを示している。 Figure 4 shows the results obtained. Since the degradation of the antibody appeared in a band around a molecular weight of 116 kilodaltons, the enlarged band is shown in the figure. Note that the darker the color of the band, the more advanced the deterioration.
　図４に示す結果より以下のことが明らかとなった。
（１）滅菌線量の影響
　放射線量依存的に抗体の劣化が引き起こされる。特に、１箇月保存した後の抗体は脱酸素剤を同封していない検体において、滅菌線量が高いほど著しい劣化を示した。
（２）滅菌線種の影響
　ガンマ線では抗体の劣化速度が非常に速いのに対し、同一量の電子線では抗体の劣化速度が遅い。ただし、電子線においても、完全には抗体の劣化を抑制できない。
（３）脱酸素剤の効果
　滅菌時、包装中に脱酸素剤を同封することにより、劣化がほぼ完全に抑制された。 The following is clear from the results shown in FIG.
(1) Effects of sterilization dose Antibody degradation is caused in a dose-dependent manner. In particular, the antibody after storage for 1 month showed a marked deterioration as the sterilization dose increased in the specimen not encapsulating the oxygen scavenger.
(2) Effect of sterilization line type While gamma rays have a very fast antibody deterioration rate, the same amount of electron beam has a slow antibody deterioration rate. However, even with an electron beam, antibody degradation cannot be completely suppressed.
(3) Effect of oxygen scavenger At the time of sterilization, the oxygen scavenger was enclosed in the package, so that the deterioration was almost completely suppressed.
　本出願は、２０１１年１０月１９日出願の米国仮出願６１／５４８，８９１に基づくものであり、その内容はここに参照として取り込まれる。 This application is based on US Provisional Application 61 / 548,891 filed on Oct. 19, 2011, the contents of which are incorporated herein by reference.
１０，２１　シングルユースバッグ
１１，２２　バッグ
１２　ポート
１３，２３　チューブ
２４，２６　包材
２５　脱酸素剤 10, 21 Single use bag 11, 22 Bag 12 Port 13, 23 Tube 24, 26 Packaging material 25 Oxygen absorber

Claims (12)

  1. 　保存容器を脱酸素雰囲気の包材中で放射線滅菌することを特徴とするたん白質の劣化を抑制する方法。 A method for suppressing protein degradation, characterized by radiation sterilization of the storage container in a packaging material in a deoxygenated atmosphere.
  2. 　前記包材中に脱酸素剤を同封することにより、該包材中を脱酸素雰囲気とする請求項１に記載の方法。 The method according to claim 1, wherein a deoxidizing atmosphere is provided in the packaging material by enclosing an oxygen scavenger in the packaging material.
  3. 　前記包材として酸素吸入性素材を用いて該包材中を脱酸素雰囲気とする請求項１または２に記載の方法。 The method according to claim 1 or 2, wherein an oxygen-absorbing material is used as the packaging material to create a deoxygenated atmosphere in the packaging material.
  4. 　脱酸素剤を同封した包材中で保存容器を放射線滅菌することを特徴とするたん白質の劣化を抑制する方法。 A method for suppressing protein degradation, characterized by radiation sterilization of a storage container in a packaging material containing an oxygen scavenger.
  5. 　前記保存容器がポリマーで構成される請求項１～４のいずれか１項に記載の方法。 The method according to any one of claims 1 to 4, wherein the storage container is composed of a polymer.
  6. 　前記ポリマーがポリエチレンである請求項５に記載の方法 The method according to claim 5, wherein the polymer is polyethylene.
  7. 　前記ポリエチレンが超低密度ポリエチレンである請求項６に記載の方法 The method according to claim 6, wherein the polyethylene is ultra-low density polyethylene.
  8. 　前記保存容器がシングルユースバッグである請求項１～７のいずれか１項に記載の方法 The method according to any one of claims 1 to 7, wherein the storage container is a single use bag.
  9. 　前記放射線滅菌がγ線滅菌である請求項１～８のいずれか１項に記載の方法。 The method according to any one of claims 1 to 8, wherein the radiation sterilization is γ-ray sterilization.
  10. 　前記たん白質が抗体である請求項１～９のいずれか１項に記載の方法。 The method according to any one of claims 1 to 9, wherein the protein is an antibody.
  11. 　前記たん白質を含む溶液を前記放射線滅菌後の前記保存容器に充填して保存する請求項１～１０のいずれか１項に記載の方法。 The method according to any one of claims 1 to 10, wherein the solution containing the protein is filled and stored in the storage container after the radiation sterilization.
  12. 　請求項１～１１のいずれか１項に記載の方法を用いてたん白質の劣化を抑制することを特徴とするたん白質を製造する方法。 A method for producing a protein, comprising suppressing deterioration of the protein using the method according to any one of claims 1 to 11.
PCT/JP2012/077140 2011-10-19 2012-10-19 Method for preventing deterioration of protein, and method for producing protein WO2013058373A1 (en)

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US61/548,891 2011-10-19

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Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1995033651A1 (en) * 1994-06-08 1995-12-14 Pharmacia & Upjohn Ab A PROCESS FOR STERILISATION BY η-RADIATION AND BY THE USE OF AN OXYGEN ABSORBER, A CONTAINER AND A MEDICAL ARTICLE STERILISED BY THE PROCESS
JPH0889561A (en) * 1994-09-22 1996-04-09 Mitsubishi Gas Chem Co Inc Sterilizing method for plastics-containing articles by irradiation with radioactive ray or electron beam
WO1997039952A1 (en) * 1996-04-23 1997-10-30 Pharmacia & Upjohn Ab Improved medical containers
JP2004275616A (en) * 2003-03-18 2004-10-07 Terumo Corp Sterilization method and method for producing medical container and prefilled syringe
WO2005004902A1 (en) * 2003-07-10 2005-01-20 Nipro Corporation Recombinant human serum albumin preparation accommodated in plastic container
JP2011025068A (en) * 1999-02-22 2011-02-10 Chugai Pharmaceut Co Ltd Pre-filled syringe protein solution product
WO2011026738A1 (en) * 2009-09-02 2011-03-10 Sanofi-Aventis Deutschland Gmbh Medicament container

Patent Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1995033651A1 (en) * 1994-06-08 1995-12-14 Pharmacia & Upjohn Ab A PROCESS FOR STERILISATION BY η-RADIATION AND BY THE USE OF AN OXYGEN ABSORBER, A CONTAINER AND A MEDICAL ARTICLE STERILISED BY THE PROCESS
JPH0889561A (en) * 1994-09-22 1996-04-09 Mitsubishi Gas Chem Co Inc Sterilizing method for plastics-containing articles by irradiation with radioactive ray or electron beam
WO1997039952A1 (en) * 1996-04-23 1997-10-30 Pharmacia & Upjohn Ab Improved medical containers
JP2011025068A (en) * 1999-02-22 2011-02-10 Chugai Pharmaceut Co Ltd Pre-filled syringe protein solution product
JP2004275616A (en) * 2003-03-18 2004-10-07 Terumo Corp Sterilization method and method for producing medical container and prefilled syringe
WO2005004902A1 (en) * 2003-07-10 2005-01-20 Nipro Corporation Recombinant human serum albumin preparation accommodated in plastic container
WO2011026738A1 (en) * 2009-09-02 2011-03-10 Sanofi-Aventis Deutschland Gmbh Medicament container

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